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目的:揭示转基因水稻全基因组遗传变异的特征与频率。创新点:通过单核苷酸分辨率揭示了农杆菌介导法转化水稻植株全基因组水平遗传变异的类型和频率以及外源DNA的整合模式。方法:应用Illumina Hiseq2000高通量测序技术测定了5个T0代转基因水稻株系的基因组序列。结合生物信息学分析和聚合酶链反应(PCR)扩增,以及Sanger测序,我们检测和验证单核苷酸多态性(SNP)和Indel变化类型和数量,转移DNA(T-DNA)及其载体骨架序列和转座子整合位点及特征,大片段的结构变异等遗传变异。结论:结果表明,农杆菌介导的水稻遗传转化,除T-DNA整合到水稻基因组外,还存在载体骨架整合现象;每个转基因水稻基因组的变异(SNP和小片段的缺失插入)数目为338–1774,与报道的组培过程中产生的变异类似;转基因水稻基因组仅存在Tos17转座子数量的变化,未检测到其他转座子数目和位置的变化。
Objective: To reveal the characteristics and frequency of genome-wide genetic variation in transgenic rice. Innovative point: Through the single nucleotide resolution revealed Agrobacterium mediated transformation of rice plant genome-wide genetic variation types and frequencies and integration of exogenous DNA patterns. METHODS: The genomic sequences of five T0 transgenic rice lines were determined using Illumina Hiseq 2000 high-throughput sequencing. Combined with bioinformatics analysis and polymerase chain reaction (PCR) amplification and Sanger sequencing, we examined and validated single nucleotide polymorphisms (SNPs) and Indel types and numbers, T-DNA and its Vector backbone sequence and transposon integration sites and features, large fragments of structural variation and other genetic variations. Conclusion: Agrobacterium-mediated rice genetic transformation, in addition to T-DNA integration into the rice genome, there is also the integration of the vector backbone; each transgenic rice genome variation (SNP and small fragment deletion insert) the number of 338 -1774, which is similar to the reported variation during tissue culture. There was only a change in the number of transposon Tos17 in the transgenic rice genome, and no change in the number and location of other transposons was detected.